A variety of analytical procedures have been developed for the separation and identification of different molecular species present in a specimen. Separation is generally accomplished by applying the specimen to a water-containing solid medium and inducing molecular separation of the species within the medium. In particular, chromatography and electrophoresis have been employed, both of which provide separation of different molecular species. The separation medium is often called a chromatographic medium or electrophoretic plate. In such processes, a variety of reagents which interact with one or more analytes in the specimen may also be applied to the medium before, during or after the separation process to assist in separation or identification of the analytes.
The detection of analytes with labeled reagents (such as labeled probes or antibodies) is a commonly used method in research and clinical laboratories. Various labels have been used, including magnetic particles, radioisotopes and enzymes, with horseradish peroxidase being one of the most common labels. Transparent, removable elements useful for producing a colorimetric signal in response to multiple analytes labeled with peroxidase are described in U.S. Pat. No. 5,047,322 (Emmons et al) and are available from International Biotechnologies, Inc. as ENZYGRAPHIC.TM. WEB.
While these elements have provided considerable advantages over the use of radioisotopes or magnetic particles as labels, there is a need for further improvement in sensitivity and a means to provide a permanent record of generated signal. The use of colorimetric signals may be suitable for immediate evaluation, but dyes tend to fade and prohibit the formation of a relatively permanent record.
It is believed that chemiluminescent assay systems would provide increased sensitivity, but current systems have several disadvantages. They require the use of wet chemistry reagents to promote the enzymatic reaction. Wet chemistry may also require dilutions of reagents, thereby reducing the sensitivity of the assay, and the handling of potentially hazardous chemicals.
U.S. Pat. No. 4,231,754 (Vogelhut) describes chemiluminescent analytical devices having multiple reagent layers which are separated from each other, for example, to form a pH gradient between layers. This is done because certain reagents are stable at relatively lower pH while a pH above 8 is desirable for effective production of chemiluminescent signal. Keeping layers at different pH values is very difficult when fluid samples or wet blot membranes are applied to the device unless immobilized buffers are used. However, having such immobilized reagents limits the usefulness of the device because only certain binders could be used in the devices. It is not believed that the devices described in this patent are practical.
More recently, U.S. Pat. No. 4,808,529 (Doppelfeld et al) describes test devices having enzymes attached directly to porous membranes having appropriate reactive groups. The signal is transferred to a transblotting membrane rather than the reaction taking place in the test device itself.
It was considered that a dry analytical element should be designed to provide chemiluminescent signals from multiple analytes. It would also be desired that such signals could be recorded in a permanent fashion since the signals do not last very long. Moreover, it would be desirable to have non-tacky, removable elements which would generate chemiluminescent signals in response to peroxidase labeled gel plates or transblotting membranes which allow the labeled plates or membranes to be used more than once. By "non-tacky" is meant that the element will not adhere to a gel plate or transblotting membrane, and can therefore be easily removed without damage to the gel plate or transblotting membrane.
It would also be desirable to have an element which has stable reagents for long term storage, is easy for the consumer to use and requires no reagent formulation.